Interleukin-10 (IL-10), a recently discovered lymphokine, was originally described as an inhibitor of interferon-.gamma. synthesis and is postulated as a major mediator of the humoral class of immune response [Fiorentino, D. F. , et al., J. Exp. Med. 170: 2081 (1989) and Moore et al., K. W. , et al., Science 248: 1230-1234 (1990)]. Two classes of often mutually exclusive immune responses are the humoral (antibody-mediated) and the delayed-type hypersensitivity.
It is postulated that these two differing immune responses may arise from two types of helper T-cell clones, namely Th1 and Th2 helper T-cells, which demonstrate distinct cytokine secretion patterns [Moore supra; Vieira, P. et al., Proc, Nat. Acad. Sci. USA Vol. 88: 1172 (1991)]. Mouse Th1 cell clones secrete interferon-.gamma. and IL-2 and preferentially induce the delayed-type hypersensitivity response while Th-2 cell clones secrete Il-4, IL-5 and IL-10 and provide support for the humoral responses [Fiorentino et al., supra]. The contrast in immune response could result because interferon-.gamma. secreted by the Th1 cell clones inhibits Th2 clone proliferation in vitro, while IL-10 secreted by the Th2 cell clones inhibit cytokine secretion by the Th1 cell clones [Fiorentino et al., supra and Moore et al., supra]. Thus the two T-helper cell types may be mutually inhibitory and may provide the underpinning for the two dissimilar immune responses.
IL-10 has been cloned and sequenced from both murine and human T cells [Moore et al., supra; Vieira et al., supra]. Both sequences contain an open reading frame encoding a polypeptide of 178 amino acids with an N-terminal hydrophobic leader sequence of 18 amino acids, and have an amino acid sequence homology of 73%.
Biologically active IL-10 is a dimer comprised of non-covalently bonded polypeptides. N-terminal analysis indicates that a small percentage of Il-10 polypeptides have the first two N-terminal amino acid residues missing. This truncated polypeptide is referred to as the .DELTA.2 IL-10 polypeptide, or simple .DELTA.2. The full-length chain is therefore referred to as .DELTA.0, indicating that no amino acid has been deleted. Accordingly, biologically active IL-10 can be expressed as three different dimers. The first biologically active dimer and the major form is .DELTA.0: .DELTA.0, a homodimer in which both polypeptides of the dimer have the full-length chain of amino acids. The second IL-10 dimer is .DELTA.0:.DELTA.2, a heterodimer in which one of the polypeptide chains has the full-length chain of amino acids and the second chain, .DELTA.2, has the first two N-terminal amino acids missing. The third IL-10 dimer is .DELTA.2:.DELTA.2, a homodimer in which both polypeptide chains of the dimer have the initial two N-terminal amino acid residues missing.
In light of its role as a potential immune response mediator and its activity as an inhibitor of interferon-.gamma. synthesis, IL-10 may have clinical utility in autoimmune diseases or transplant rejection. However, in a clinical setting it is highly desirable that the IL-10 be in a highly pure state, substantially free of other contaminating host and medium proteins or polypeptides.
Thus there is a need for purifying IL-10 from culture medium, and in particular there is a need for a method for separating the different dimers of IL-10 from each other.